Cervical cancer, like many other cancers, metabolizes glucose primarily through aerobic glycolysis, i.e., the Warburg effect, and this altered tumor cell physiology can be imaged by 18F-fluorodeoxyglucose-positron emission tomography (FDG-PET). This project is designed to test the hypothesis that tumors with increased glucose uptake on pretreatment FDG-PET imaging are inherently radio-resistant, and this radio-resistance can be reversed by taking advantage of drug combinations that specifically target tumor glucose metabolism. Preliminary data has shown that constitutive activation of PI3K/AKT signaling may be one mechanism that drives aberrant glucose metabolism in cervical cancer, and activating mutations in PIK3CA are associated with increased distant metastasis and decreased survival outcome after standard chemoradiation (pelvic irradiation plus concurrent cisplatin chemotherapy). The experiments in the current proposal are being performed to generate preclinical data in support of using PI3K/AKT pathway inhibitor combinations as radio-sensitizers for metabolically active cervical tumors. The research strategy employs a mechanistic approach to study PI3K/AKT pathway alterations in cervical carcinoma and develops a novel strategy for targeted radio-sensitization that combines an inhibitor of AKT signaling with an inhibitor of glycolysis (2-DG). The objective of Specific Aim #1 is to determine if thiol-mediated oxidative stress caused by inhibition of glucose metabolism contributes to the mechanism by which PI3K/AKT inhibition radio-sensitizes cervical cancer cells. The objective of Specific Aim #2 is to determine whether alterations in PI3K/AKT signaling (i.e., mutations and inhibitors) affect FDG uptake and the radiation response in vivo by using a novel, clinically relevant model of human cervical cancer in the mouse. These tumors can be manipulated in vitro prior to implantation to directly test the function of individual genes or to evaluate the effects of specific gene mutations, which will provide a rigorous and reproducible system to evaluate inhibitor combinations and to determine whether individual gene mutations can serve as biomarkers for response to treatment. The objective of Specific Aim #3 is to develop a metabolic signature that can be used to identify patients for treatments with targeted radio-sensitization with PI3K/AKT inhibitors plus 2-DG. Development of the metabolic signature will employ an innovative approach using a combination of patient-specific genomic and metabolomic data. If validated, the metabolic signature developed in Aim 3 could be used in the future to select patients for treatment with PI3K/AKT inhibitors +/- 2-DG as a novel approach for targeted radio-sensitization of metabolically active and radio-resistant tumors. Because alterations in the PI3K/AKT pathway and the regulation of glucose metabolism are common in human cancer, the results of this research have the potential to improve radiotherapy outcome for many other cancers including tumors of the breast, uterus, pancreas and prostate.